CN108220355A - The processing method of Production by Enzymes alpha-arbutin waste liquid and its application - Google Patents

The processing method of Production by Enzymes alpha-arbutin waste liquid and its application Download PDF

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CN108220355A
CN108220355A CN201711472244.XA CN201711472244A CN108220355A CN 108220355 A CN108220355 A CN 108220355A CN 201711472244 A CN201711472244 A CN 201711472244A CN 108220355 A CN108220355 A CN 108220355A
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waste liquid
arbutin
production
processing method
alpha
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郭恒华
张冬竹
田宋魁
刘树蓬
章晖
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Anhui Huaheng Biotechnology Co Ltd
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Anhui Huaheng Biotechnology Co Ltd
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P13/00—Preparation of nitrogen-containing organic compounds
    • C12P13/04—Alpha- or beta- amino acids
    • C12P13/14—Glutamic acid; Glutamine
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P13/00—Preparation of nitrogen-containing organic compounds
    • C12P13/04—Alpha- or beta- amino acids
    • C12P13/06—Alanine; Leucine; Isoleucine; Serine; Homoserine

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  • Organic Chemistry (AREA)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
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  • Life Sciences & Earth Sciences (AREA)
  • Wood Science & Technology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Microbiology (AREA)
  • General Chemical & Material Sciences (AREA)
  • Biotechnology (AREA)
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  • Biochemistry (AREA)
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Abstract

The present invention relates to technical field of chemistry, and in particular to a kind of processing method of Production by Enzymes α ursin waste liquids and its application；Include the following steps：(1) PH to 4~5 of α ursin production waste liquid is adjusted using acid regulator, carbohydrase is added in and reaction is hydrolyzed；(2) waste liquid PH to 3~4 is adjusted using acid regulator, flocculation, filtering adjust waste liquid PH to neutrality using alkaline conditioner later, obtain conversion fluid, conversion fluid finally is configured to fermentation medium；The processing method provided by the present invention is simple, processing cost is low, resource utilization is high.

Description

The processing method of Production by Enzymes alpha-arbutin waste liquid and its application
Technical field
The present invention relates to liquid waste disposal technique fields, and in particular to a kind of processing method of Production by Enzymes alpha-arbutin waste liquid And its application.
Background technology
Alpha-arbutin is the epimer of β-ursin, the entitled 4- hydroxyphenyl-α-D- glucopyranosides of chemistry.It grinds Study carefully and show that alpha-arbutin is more stable compared with property for β-ursin, there is preferably anti-inflammatory, reparation and white-skinned face function, frequently as Brightening agent is applied in high-end skin care item and cosmetics.
In the prior art, the main production process of alpha-arbutin is Enzyme optrode, is produced using in microbial cell Enzyme, be catalyzed and polysaccharide and form single alpha-arbutin with reacted with hydroquinone.In process of production, it is extracted through process for separating and purifying After alpha-arbutin, there are the compounding substances such as polysaccharide, monosaccharide, surfactant in production waste liquid to be difficult to handle.At present, for the life Waste liquid is produced, there are two types of common processing methods：First, arrange as conventional sewage is outer after processing, not only waste of resource in this way, also Increase processing cost.Second is that saccharomycete is added in into waste liquid removes thalline to remove glucose, then by membrane filtration, due to outer Purchase saccharomycete needs to increase production cost, and the metabolite complicated component that yeast generates during the fermentation, membrane filtration processing Difficulty is big, of high cost.
Invention content
The purpose of the present invention is to provide the Production by Enzymes α that a kind of method is simple, processing cost is low, resource utilization is high- The processing method of ursin waste liquid.
To achieve the above object, the technical solution adopted by the present invention is：A kind of processing of Production by Enzymes alpha-arbutin waste liquid Method includes the following steps：
(1) PH to 4~5 of alpha-arbutin production waste liquid is adjusted using acid regulator, carbohydrase is added in and is hydrolyzed instead It should；
(2) waste liquid PH to 3~4 is adjusted using acid regulator, flocculation, filtering are adjusted useless using alkaline conditioner later Liquid PH obtains conversion fluid, conversion fluid finally is configured to fermentation medium to neutrality.
In the above-mentioned technical solutions, pH is first adjusted to 4~5, at this time the enzyme activity highest of carbohydrase, sucrose can most efficiently, It is converted to glucose and fructose to the greatest extent.Then pH is adjusted to 3~4, maintains a period of time, flocculate to albumen, To remove deproteinized.PH is finally adjusted to neutrality, can guarantee after culture medium is configured to the conversion fluid, culture can be supplied to The suitable growing environment of strain in base.Our experiments show that the ingredient of pending alpha-arbutin production waste liquid is：Sucrose 170g/ L, fructose 144g/L, surfactant 0.1%, pH6.5~7.5.After the above method is handled, conversion fluid ingredient is：Glucose 90g/L, fructose 234g/L, also surfactant 0.1%, NaCl or Na2SO4.It is prolonged through applicant the study found that Can be used for preparing all kinds of fermentation mediums through the method treated waste water, this is because glucose, fructose can be used as carbon source and The energy is utilized by microorganism, NaCl or Na2SO4Inorganic salts necessary to be microorganism growth, surfactant can increase cell The permeability of film reduces the transmission resistance into cell of oxygen and nutriment, increases the release of tunning, improves yield. The processing method provided by the present invention is simple, processing cost is low, can realize the effective recycling of resource.
Description of the drawings
Fig. 1 is the culture medium (A) that Application Example 1 is prepared and the growth of culture medium (B) that comparative example 1 is prepared Rate comparison figure；
Fig. 2 is the culture medium (A) that Application Example 1 is prepared and the l-Alanine of culture medium (B) that comparative example 1 is prepared Accumulation rate comparison diagram；
Fig. 3 is the culture medium (C) that Application Example 2 is prepared and the growth of culture medium (D) that comparative example 2 is prepared Rate comparison figure；
Fig. 4 is the culture medium (C) that Application Example 1 is prepared and the Pidolidone of culture medium (D) that comparative example 1 is prepared Accumulation rate comparison diagram.
Specific embodiment
1-3 is further described technical solution disclosed by the invention with reference to embodiments.
Embodiment 1：Alpha-arbutin produces the processing of waste liquid
(1) using 0.5mol/LH2SO4Solution adjusts alpha-arbutin production waste liquid PH to 4.5, adds in carbohydrase, and reaction 2 is small When；The alpha-arbutin produces component of effluent：Sucrose 170g/L, fructose 144g/L, surfactant 0.1%；Waste liquid PH6.8, volume are 5.0L；
(2) using 0.5mol/LH2SO4Waste liquid PH to 3.5 is adjusted, after 5 minutes, flocculation, filtering, then using NaOH tune Waste liquid PH is saved to neutrality, obtains conversion fluid.After processing, conversion fluid ingredient is：Glucose about 90g/L, fructose about 234g/L, table Face activating agent about 0.1%, micro Na2SO4。
Embodiment 2：Alpha-arbutin produces the processing of waste liquid
(1) PH to 4 of alpha-arbutin production waste liquid is adjusted using 0.5mol/LHCl, carbohydrase is added in, reacts 1 hour；
(2) waste liquid PH to 3 is adjusted using 0.5mol/LHCl, after 5 minutes, then flocculation, filtering are adjusted using NaOH Waste liquid PH obtains conversion fluid to neutrality.
Embodiment 3：Alpha-arbutin produces the processing of waste liquid
(1) PH to 5 of alpha-arbutin production waste liquid is adjusted using 0.5mol/LHCl, carbohydrase is added in, reacts 2 hours；
(2) using 0.5mol/LH2SO4Waste liquor PH is adjusted to after 4,5 minutes, then flocculation, filtering are adjusted using NaOH Waste liquor PH obtains conversion fluid to neutrality.
Application Example 1：The preparation of l-Alanine fermentation medium
(1) conversion fluid that 1 processing of 0.39L embodiments obtains is measured；
(2) following substance is added into conversion waste liquid：Ammonium chloride 5g, NaH2PO45g, Na2HPO45g, MgSO4·7H2O 1g, CaCl2·2H2O 0.1g, small-scale inorganic salt 2.1mg, small-scale inorganic salt composition are：FeCl3·6H2O 1.5mg, CoCl2· 6H2O 0.1mg, CuCl2·2H2O 0.1mg, ZnCl20.1mg, Na2MoO4·2H2O 0.1mg, MnCl2·4H2O2 0.2mg。 Distilled water is supplemented to 1L, it is 6.5 to adjust medium pH, obtains l-Alanine fermentation medium.
Ingredient includes in the l-Alanine fermentation medium：Sugar (glucose and fructose) 100g/L, ammonium chloride 5g/L, NaH2PO45g/L, Na2HPO45g/L, MgSO4·7H2O 1g/L, CaCl2·2H2O 0.1g/L, surfactant is about 0.04%, small-scale inorganic salt 2.1mg/L, small-scale inorganic salt composition are：FeCl3·6H2O 1.5mg, CoCl2·6H2O 0.1mg, CuCl2·2H2O 0.1mg, ZnCl20.1mg, Na2MoO4·2H2O 0.1mg, MnCl2·4H2O20.2mg, culture medium pH6.5。
Comparative example 1：The preparation of normal fermentation culture medium 1
Normal fermentation culture medium composition is as follows：Glucose 100g/L, ammonium chloride 5g/L, NaH2PO45g/L, Na2HPO4 5g/L, MgSO4·7H2O 1g/L, CaCl2·2H2O 0.1g/L, small-scale inorganic salt 2.1mg/L, medium pH 6.5.Micro nothing Machine salt forms：FeCl3·6H2O 1.5mg, CoCl2·6H2O 0.1mg, CuCl2·2H2O 0.1mg, ZnCl20.1mg, Na2MoO4·2H2O 0.1mg, MnCl2·4H2O2 0.2mg。
Application Example 2：The preparation of Pidolidone fermentation medium
(1) conversion fluid that 1 processing of 0.463L embodiments obtains is measured；
(2) following substance is added into conversion waste liquid：20g NH4Cl, 45g CaCl2, 1gKH2PO4, 1g peptones, 400mg MgSO4·7H2O, 10mgFeSO4·7H2O, 10mg MnSO4·7H2O, 300 μ g biotins, 50 μ g thiamine hydrochlorides, With 4mg chloramphenicol, supplement distilled water to 1L, it is 7.8 to adjust medium pH, obtains Pidolidone fermentation medium.
Ingredient includes in Pidolidone fermentation medium：Sugar (glucose and fructose) 150g/L, 20g/L NH4Cl, 45g/L CaCl2, 1g/L KH2PO4, 1g/L peptones, 400mg/L MgSO4·7H2O, 10mg/L FeSO4·7H2O, 10mg/L MnSO4·7H2O, 300 μ g/L biotins, 50 μ g/L thiamine hydrochlorides and 4mg/L chloramphenicol, surfactant concentration are 0.0463%, medium pH 7.8.
Comparative example 2：The preparation of normal fermentation culture medium 2
The composition of normal fermentation culture medium 2 is as follows：Glucose 150g/L, 20g/L NH4Cl, 45g/L CaCl2, 1g/L KH2PO4, 1g/L peptones, 400mg/L MgSO4·7H2O, 10mg/L FeSO4·7H2O, 10mg/L MnSO4·7H2O, 300 μ G/L biotins, 50 μ g/L thiamine hydrochlorides and 4mg/L chloramphenicol, medium pH 7.8.
Performance test is tested：
First, under same experiment condition, using the l-Alanine fermented and cultured in Application Example 1 and comparative example 1 Base and normal fermentation culture medium 1 carry out fermented and cultured to the bacterial strain of l-Alanine (CGMCC No.4036) respectively, and fermentation temperature is 35 DEG C, pH 6.5, speed of agitator 500rpm, ventilation quantity 600m3/ h, fermentation results are to such as Tables 1 and 2, Fig. 1 and Fig. 2 institutes Show：
Table 1:Growth rate comparison (OD)
Detection moment (h) 0 1 2 3 4 5 6 7 8 9 10
A 0.00 0.02 0.12 0.29 0.43 0.89 1.34 2.69 4.12 7.89 16.60
B 0.00 0.02 0.09 0.26 0.46 0.96 1.29 2.12 4.89 7.56 13.53
Detection moment (h) 11 12 13 14 15 16 17 18 19 20
A 23.60 24.50 23.20 22.30 21.40 20.90 21.10 20.10 19.81 19.23
B 21.63 22.65 25.10 24.53 23.63 22.15 21.11 19.63 19.79 20.13
Table 2：L-Alanine accumulation rate compares (g/L)
Detection moment (h) 0 1 2 3 4 5 6 7 8 9 10
A 0 0.11 0.24 0.31 0.36 0.41 0.81 1.11 1.23 1.76 2.15
B 0 0.09 0.23 0.39 0.31 0.45 0.89 1.23 1.19 1.56 1.98
Detection moment (h) 11 12 13 14 15 16 17 18 19 20
A 3.26 5.36 7.36 17.5 21.3 32.9 38.3 46.3 51.2 64.3
B 2.96 4.63 6.98 15.2 18.6 30.3 42.3 44.2 48.2 67.9
It should be noted that in table 1, table 2, A is the culture medium that Application Example 1 is prepared；B is prepared for comparative example 1 Culture medium.
From Tables 1 and 2, Fig. 1 and Fig. 2 can be seen that：The l-Alanine fermentation medium and routine be configured to by conversion fluid Fermentation medium 1 compares, and the growth rate of strain and the accumulation rate of l-Alanine are consistent, shows using made of conversion fluid Culture medium has good application effect.
2nd, under same experiment condition, using the Pidolidone fermented and cultured in Application Example 2 and comparative example 2 Base and normal fermentation culture medium 2 respectively carry out brevibacterium flavum fermented and cultured, and fermentation temperature is 35 DEG C, pH 7.8, and stirring turns Speed is 500rpm, ventilation quantity 600m3/ h, fermentation results are to such as table 3 and table 4, shown in Fig. 3 and Fig. 4：
Table 3：Growth rate comparison (OD)
Table 4：Pidolidone accumulation rate compares (g/L)
Detection moment (h) 0 1 2 3 4 5 6 7 8 9 10
A 0 1.95 2.16 3.26 3.35 5.39 6.27 7.34 9.62 10.67 14.53
B 0 1.85 1.96 2.98 3.05 4.56 5.68 6.64 8.88 10.23 13.24
Detection moment (h) 11 12 13 14 15 16 17 18 19 20
A 17.48 25.12 32.99 39.52 51.89 59.64 69.61 74.69 81.54 86.46
B 18.56 22.31 28.67 41.81 45.21 62.68 67.23 74.69 83.29 83.1
It should be noted that in table 3, table 4, C is the culture medium that Application Example 2 is prepared；D is prepared for comparative example 2 Culture medium.
It can be seen that from table 3,4, Fig. 3,4：Compared by the Pidolidone that conversion fluid is configured to and normal fermentation culture medium 2 Compared with the growth rate of strain and the accumulation rate of Pidolidone are consistent, show have very well using culture medium made of conversion fluid Application effect.

Claims (8)

1. a kind of processing method of Production by Enzymes alpha-arbutin waste liquid, which is characterized in that include the following steps：
(1) PH to 4~5 of alpha-arbutin production waste liquid is adjusted using acid regulator, carbohydrase is added in and reaction is hydrolyzed；
(2) waste liquid PH to 3~4 is adjusted using acid regulator, flocculation, filtering adjust waste liquid PH using alkaline conditioner later To neutrality, conversion fluid is obtained, conversion fluid is finally configured to fermentation medium.
2. the processing method of Production by Enzymes alpha-arbutin waste liquid according to claim 1, which is characterized in that the acid tune Section agent is HCl or H2SO4。
3. the processing method of Production by Enzymes alpha-arbutin waste liquid according to claim 1, which is characterized in that the alkalinity is adjusted Section agent is NaOH.
4. the processing method of Production by Enzymes alpha-arbutin waste liquid according to claim 1, which is characterized in that the step (1) time of hydrolysis is 1-3 hours in.
What 5. a kind of processing method of Production by Enzymes alpha-arbutin waste liquid according to any one of claim 1-4 obtained turns Change the application of liquid in the fermentation medium.
6. application according to claim 5, which is characterized in that the fermentation medium includes l-Alanine fermented and cultured Base, Pidolidone fermentation medium.
7. application according to claim 6, which is characterized in that contain in every liter of l-Alanine fermentation medium：Portugal Grape sugar+90~110g of fructose, ammonium chloride 3~6g, NaH2PO43~6g, Na2HPO43~6g, MgSO4·7H2O0.5~2g, CaCl2·2H20.05~0.2g of O, 1.5~3mg of small-scale inorganic salt, surfactant concentration are 0.01~0.07%；It is described micro- Amount inorganic salts, which form, is：FeCl3·6H2O 0.5~2mg, CoCl2·6H2O 0.05~1.5mg, CuCl2·2H2O 0.05~ 0.2mg, ZnCl20.05~2mg, Na2MoO4·2H2O 0.05~2mg, MnCl2·4H2O20.1~0.3mg.
8. application according to claim 6, which is characterized in that contain in every liter of Pidolidone fermentation medium：Portugal Grape sugar+fructose 120~170g, NH4Cl 15~25g, CaCl230~50g, KH2PO40.5~2g, 0.5~2g of peptone, MgSO4·7H2O 350~450mg, FeSO4·7H2O 5~15mg, MnSO4·7H25~15mg of O, 250~350 μ of biotin G, 30~60 μ g of thiamine hydrochloride, 1~6mg of chloramphenicol, surfactant concentration is 0.01~0.06%.
CN201711472244.XA 2017-12-29 2017-12-29 The processing method of Production by Enzymes alpha-arbutin waste liquid and its application Pending CN108220355A (en)

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Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1635139A (en) * 2004-11-15 2005-07-06 刘春巧 Process for preparing alpha-arbutin through fermentation
CN102978265A (en) * 2012-12-03 2013-03-20 韦慧芳 Method for synthesizing alpha-arbutin by enzymic method through catalysis
CN105400851A (en) * 2015-12-25 2016-03-16 天津宏顺科生物科技有限公司 Preparation method of alpha-arbutin

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1635139A (en) * 2004-11-15 2005-07-06 刘春巧 Process for preparing alpha-arbutin through fermentation
CN102978265A (en) * 2012-12-03 2013-03-20 韦慧芳 Method for synthesizing alpha-arbutin by enzymic method through catalysis
CN105400851A (en) * 2015-12-25 2016-03-16 天津宏顺科生物科技有限公司 Preparation method of alpha-arbutin

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